CLONING, EXPRESSION, AND FUNCTIONAL-CHARACTERIZATION OF 2 MUTANT (NAT2(191) AND NAT2(341 803)) AND WILD-TYPE HUMAN POLYMORPHIC N-ACETYLTRANSFERASE (NAT2) ALLELES/

The N-acetylation polymorphism segregates individuals into rapid, inte rmediate, and slow acetylator phenotypes via monogenic inheritance at the NAT2 locus. In a previous study (Arch. Toxicol. 67, 445-452, 1993) , we uncovered discrepancies between apparent NAT2 acetylator genotype based on polymerase chain reaction-restriction fragment length polymo rphism analysis, in vitro colon arylamine N-acetyltransferase activity , and expected frequency of slow acetylator phenotype in African-Ameri cans, which suggested the presence of not yet defined mutant NAT2 alle les. Two novel NAT2 alleles were discovered after cloning and sequenci ng of NAT2 polymerase chain reaction products. One allele (NAT2(191)) contained a point mutation at nucleotide 191 [G --> A (Arg --> Gln)], whereas the other allele (NAT2(341/803)) contained two point mutations [341T --> C (lle --> Thr); 803A --> G (Lys --> Arg)]. The two mutant NAT2 and the NAT2(wt) alleles were expressed in a prokaryotic expressi on system. Both the NAT2(191) and NAT2(341/803) mutant alleles express ed functional N-acetyltransferases capable of catalyzing both arylamin e N-acetylation and the metabolic activation (via O-acetylation) of N- hydroxy-2-aminofluorene. However, the NAT2(191) and NAT2(341/803) each exhibited significantly lower N- and O-acetylation capacity and were intrinsically less stable than NAT2(wt).